Ly6g pe 1a8

Manufactured by BioLegend

Sourced in United States

Ly6G-PE (1A8) is a monoclonal antibody that binds to the Ly6G antigen, a cell surface marker expressed on granulocytes such as neutrophils. This antibody is conjugated with the fluorescent dye Phycoerythrin (PE) for detection and analysis purposes.

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Lab products found in correlation

Apc gr1 rb6 8c5, by BioLegend (2 mentions) Hts transwell 96 well plate, by Corning (1 mentions) Tcs sp5, by Leica (1 mentions) Cd11b fitc m1 70, by BD (1 mentions) Sc30 digital camera, by Olympus (1 mentions) Axioplan microscope, by Olympus (1 mentions) Gl7 fitc, by BD (1 mentions) Cd138 pe 281 2, by BD (1 mentions) B220 apc ra3 6b2, by BD (1 mentions) Lsr 2 flow cytometer, by BD (1 mentions) F4 80 pe cy7 bm8, by BioLegend (1 mentions) Foxp3 transcription factor staining kit, by Thermo Fisher Scientific (1 mentions) Fixable viability stain 520, by BD (1 mentions) Cd3 percp cy5.5 17a2, by BioLegend (1 mentions) Fortessa flow cytometer, by BD (1 mentions) Ly6c apc hk1, by BioLegend (1 mentions) Cd8a pe 53 6, by BioLegend (1 mentions) Cd4 apc gk1.5, by BioLegend (1 mentions) Cd25 apc 3c7, by BioLegend (1 mentions) Nk1.1 apc pk136, by BD (1 mentions)

Spelling variants of this product

Ly6G (1A8, (66 citations) Ly6G-PE (49 citations) Anti–Ly6G-PE (clone 1A8; (5 citations) FITC-conjugated or PE-Cy7-conjugated anti-Ly6G (clone 1A8, (4 citations) Ly6G-PE (clone 1A8) (3 citations) PE)-conjugated anti-mouse Ly6G antibody (clone 1A8), (2 citations)

7 protocols using ly6g pe 1a8

Neutrophil Migration and Transendothelial Assays

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Migration assays were performed in Zigmond chambers according to the manufacturer. In brief, Ly6G-PE (1A8, BioLegend) labelled neutrophils were seeded (30 min) on a collagen-coated coverslip and mounted on a glass slide chamber. A gradient of PDGF-BB activated SMC vs non-activated SMC supernatants was created and images were acquired over 30 min in 30 s intervals using a climate chamber fluorescence microscope (20x dry objective, Leica, DMi8). Speed and displacement were calculated as previously described^{20 (link)}.
To analyse transmigration, neutrophils (2 × 10⁵) were added to the top compartment of HTS transwell 96 well plates (Corning) with a 3-μm pore size. In the lower compartment, supernatants obtained from non-activated or PDGF-BB-activated SMCs were added. After incubation for 1 h at 37 °C, transmigrated neutrophils were analysed from the bottom compartment by flow cytometry.

Silvestre-Roig C., Braster Q., Wichapong K., Lee E.Y., Teulon J.M., Berrebeh N., Winter J., Adrover J.M., Santos G.S., Froese A., Lemnitzer P., Ortega-Gómez A., Chevre R., Marschner J., Schumski A., Winter C., Perez-Olivares L., Pan C., Paulin N., Schoufour T., Hartwig H., González-Ramos S., Kamp F., Megens R.T., Mowen K.A., Gunzer M., Maegdefessel L., Hackeng T., Lutgens E., Daemen M., von Blume J., Anders H.J., Nikolaev V.O., Pellequer J.L., Weber C., Hidalgo A., Nicolaes G.A., Wong G.C, & Soehnlein O. (2019). Externalized histone H4 orchestrates chronic inflammation by inducing lytic cell death. Nature, 569(7755), 236-240.

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Multicolor Immunofluorescence Staining of Splenic Cells

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Spleens were frozen in OCT (Sakura) and cut into 8-μm-thick sections using a cryostat microtome. The sections were dried overnight, blocked with 5% goat sera (Dako), and stained with the following anti-mouse antibodies: B220-APC (RA3-6B2; BD Biosciences), CD138-PE (281-2; BD Biosciences), GL-7-FITC (BD Biosciences), Ly6G-PE (1A8; BioLegend), hCD2-FITC (RPA-2.10, BioLegend), IgG3-APC (R40-82), and CD11b-FITC (M1/70; BD Biosciences). Images were acquired on a confocal laser scanning microscope (Leica; TCS SP5) and processed with Photoshop software (Adobe Systems). For histopathological evaluation of the joints, paws were removed from the mice at the end point (day 61) and fixed by immersion in 4% paraformaldehyde. Paws were then decalcified in 10% ethylenediaminetetraacetate for 2 to 3 wk at room temperature, followed by paraffin embedding and sectioning. Deparaffinized and rehydrated slides were stained with H&E and used to assess joint inflammation and bone degradation (seven slides per joint at 5- and 200-μm intervals between slides). Images were acquired on a Zeiss Axioplan microscope that was equipped with an Olympus SC30 digital camera.

Sedimbi S.K., Hägglöf T., Garimella M.G., Wang S., Duhlin A., Coelho A., Ingelshed K., Mondoc E., Malin S.G., Holmdahl R., Lane D.P., Leadbetter E.A, & Karlsson M.C. (2020). Combined proinflammatory cytokine and cognate activation of invariant natural killer T cells enhances anti-DNA antibody responses. Proceedings of the National Academy of Sciences of the United States of America, 117(16), 9054-9063.

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Cardiac Tissue Immune Cell Analysis

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Cardiac tissue was isolated after sham or TAC surgery. Tissue was digested with collagenase I (450 U/mL), collagenase XI (125 U/mL), DNase I (60 U/mL), and hyaluronidase (60 U/mL), and the isolated cells from digested tissue were immunostained with fluorescently conjugated antibodies against CD45‐Pacific Blue, 30‐F11 (BioLegend Catalog #103126), Ly6G‐PE, 1A8 (BioLegend Catalog #127618), CD11b‐APC‐Cy7, M1/70 (BioLegend Catalog #101226), F4/80‐PE‐Cy7, BM8 (BioLegend Catalog #123114), and Ly6C‐FITC, HK1.4 (BioLegend Catalog #128006). Dead cells were excluded by staining with DAPI. Immunostained fluorescent cells were analyzed by flow cytometry using a BD LSR II flow cytometer (BD Bioscience). Additionally, isolated cardiac tissue was lysed and analyzed for inflammatory cytokines interleukin‐1β, inrerleukin‐6, and C‐C motif chemokine ligand 2 by quantitative reverse transcription PCR. Real‐time PCR primers are listed in Table S1.

Chavkin N.W., Sano S., Wang Y., Oshima K., Ogawa H., Horitani K., Sano M., MacLauchlan S., Nelson A., Setia K., Vippa T., Watanabe Y., Saucerman J.J., Hirschi K.K., Gokce N, & Walsh K. (2021). The Cell Surface Receptors Ror1/2 Control Cardiac Myofibroblast Differentiation. Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease, 10(13), e019904.

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Flow Cytometry Analysis of Mouse Tumor Infiltrates

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Single-cell suspensions from mouse subcutaneous tumors were prepared as described above, counted, and resuspended in PBS at a concentration of 1×10⁷ live cells/mL. One hundred microliters of each sample was plated for cell staining. Surface staining was performed at room temperature for 30 min, and intracellular staining was performed using a Foxp3-transcription factor staining kit (eBioscience). Live/dead cell discrimination was performed using Fixable Viability Stain 520 (BD Biosciences). Anti-mouse antibodies against the following antigens were used for flow cytometry: CD3 PerCP-CY5.5 (17A2, BioLegend, 1:100), CD45 PerCP (30-F11, BioLegend, 1:200), NK1.1 APC (PK136, BD Biosciences, 1:50), Foxp3 PE (MF23, BD Biosciences, 1:100), Ly6G PE (1A8, BioLegend, 1:100), Ly6C APC (HK1.4, BioLegend, 1:100), CD11b BV421 (M1/70, BioLegend, 1:100), CD11b PerCP-CY5.5 (HL3, BioLegend, 1:100), F4/80 APC (T45–2342, BD Biosciences, 1:50), CD25APC (3C7, BioLegend, 1:100), CD4 APC (GK1.5, BioLegend, 1:100), and CD8a PE (53-6.7; BioLegend, 1:100). All flow cytometry analyses were performed using a Fortessa flow cytometer (BD Biosciences) and analyzed using FlowJo software (TreeStar).

Huang Q., Liu J., Wu S., Zhang X., Xiao Z., Liu Z, & Du W. (2021). Spi-B Promotes the Recruitment of Tumor-Associated Macrophages via Enhancing CCL4 Expression in Lung Cancer. Frontiers in Oncology, 11, 659131.

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Neutrophil Identification in Myocardial Infarction

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Mice were sacrificed 1 day after myocardial infarction. Peripheral blood was drawn through the cardiac puncture into spray-coated K2EDTA tubes (BD Biosciences). Red blood cells were lysed with the RBC lysis buffer (BioLegend). And the remaining cells were resuspended with PBS containing 0.5% (wt/vol) BSA. The cells were incubated with the following antibodies: anti-CD11b-APC, M1/70 (BioLegend); -Ly6C-PerCP/Cy5.5, HK1.4 (BioLegend); and -Ly6G-PE, 1A8 (BioLegend). Neutrophils were identified as CD11b⁺ Ly6G^high Ly6C^low. Data were acquired on the BD FACSAria™ II.

Zhou Z., Zhang S., Ding S., Abudupataer M., Zhang Z., Zhu X., Zhang W., Zou Y., Yang X., Ge J, & Hong T. (2019). Excessive Neutrophil Extracellular Trap Formation Aggravates Acute Myocardial Infarction Injury in Apolipoprotein E Deficiency Mice via the ROS-Dependent Pathway. Oxidative Medicine and Cellular Longevity, 2019, 1209307.

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Autophagy Modulation in Inflammation

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Ethyl pyruvate, lipopolysaccharide and N-Formyl-Met-Leu-Phe were obtained from Sigma-Aldrich (Merck Millipore, Darmstadt, Germany). Antibodies against LC3 (cat. no. 4108S, 1:2,000), Becn1 (cat. no. 3738S, 1:2,000), ATG5 (cat. no. 12994, 1:2,000), and β-actin (cat. no. 4970S, 1:2,000) were obtained from Cell Signaling Technology, Inc. (Danvers, MA, USA). PE-Ly6G [1A8] and APC-Gr1 [RB6-8C5] were purchased from BioLegend, Inc. (San Diego, CA, USA). ELISA Kits for tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and MPO were purchased from R&D Systems, Inc. (Minneapolis, MN, USA).

Zhu Q., Wang H., Wang H., Luo Y., Yu Y., Du Q., Fei A, & Pan S. (2017). Protective effects of ethyl pyruvate on lipopolysaccharide-induced acute lung injury through inhibition of autophagy in neutrophils. Molecular Medicine Reports, 15(3), 1272-1278.

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Flow Cytometric Analysis of BALF Cells

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BALF cells and lung leukocytes were assessed using flow cytometry. BAL cells (50,000 cells) in 100 µl flow assay buffer were incubated with PE-Ly6G [1A8] (#127607) and APC-Gr1 [RB6-8C5] (#108411) (Biolegend, San Diego, CA, USA). Cells were washed again, resuspended in 3% paraformaldehyde and analyzed using a FACS Caliber flow cytometer (BD Bioscience, San Jose, CA, USA).

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